Exchange of extracellular vesicles (EVs, referring to exosomes and microvesicles) has emerged as a novel mechanism of cell-cell communication in the central nervous system, but very little is known about the biological function of those transfers in health or disease. Astrocytes, one of the most abundant cell types of the central nervous system (CNS), secrete EVs that can be beneficial or detrimental to neuronal fitness depending on their activation state, thus implying that astrocyte-derived EVs can modulate brain resilience and vulnerability to stress. Interestingly, we have recently observed that apolipoprotein E (APOE), the most relevant genetic risk modulator of Alzheimer?s disease (AD), can be found in association with EVs isolated from astrocytes in vitro, the major cell type producing APOE in the brain. While APOE has been known for decades as the primary carrier of lipids between neural cells and a strong partner of Amyloid ? peptides in AD (APOE4 and APOE2 respectively increasing or decreasing amyloid buildup in the brain and conferring higher risk or protection toward the disease), this discovery suggests a novel role of APOE in modulating the transfer of many other biologically active compounds through EVs, including nucleic acids. The present proposal therefore hypothesizes that the exchange of EVs between astrocytes and neurons can be differentially modulated by each APOE isoform, depends upon astroglial reactivity and is, at least in part, responsible for some non-cell autonomous effects in aging and Alzheimer?s disease. To investigate those exciting questions, we will further characterize the association of APOE with astrocyte-derived EVs from human brain samples, analyze the nucleic acid content of EVs released from resting or reactive astrocytes and monitor in vivo and in vitro, how those parameters (APOE isoforms and activation state of astrocytes) may eventually impact EV neuronal uptake. Importantly, because EVs are small particles that cannot be readily observed by conventional microscopy and because all cell types can secrete and capture EVs, studying their exchanges in vivo has proven very challenging. To circumvent this problem, we have developed a novel reporter system based on the secretion of astrocyte-derived EVs containing Cre recombinase mRNA (donor cell type) and on the expression of a Cre-sensitive reporter in neurons (recipient cell type). Using this innovative ?ON/OFF? reporter system, we will be able, for the first time, to study the dynamic transfers of EVs from astrocytes to neurons in the living mouse brain by multiphoton-imaging, determine if those processes are modulated by the nature of each APOE variant and exacerbated after LPS-induced neuroinflammation, upon aging or in the context of AD neuropathological changes. Importantly, we will also be able to sort recipient neurons from non-recipient neurons from the same biological environment and establish the specific transcriptional changes that distinguish both populations, eventually correlating those findings with our initial screen of RNA cargos detected in EVs isolated from resting and reactive astrocytes. Considering the paucity of knowledge in the field, the current project will advance our understanding in the biological relevance of EV-based cell-cell communication in the context of aging or neurodegeneration, eventually opening novel avenues of investigation.